Role ofsipAin the early stages ofSalmonella typhimuriumentry into epithelial cells

Abstract

Salmonella virulence depends on an ability to invade host cells, which is in turn dependent on a type III protein secretion system encoded in Salmonella pathogenicity island 1 (SPI1). Several protein targets of the SPI1‐encoded secretion system are translocated into host cells, where they subvert cellular processes that contribute to bacterial invasion, actin rearrangement, membrane ruffling and other aspects of virulence. We examined the role of sipA (encoding the translocated protein SipA) and found that a sipA mutant was significantly less invasive in Madin–Darby canine kidney (MDCK) cells than in its parental strain at the earliest stages of infection (5 min). The invasion defect associated with sipA was no longer apparent after 15 min of infection. Confocal microscopy of F‐actin in tetramethyl rhodamine isothiocyanate (TRITC)–phalloidin‐stained MDCK cells revealed no difference in either the frequency or the morphology of membrane ruffles induced by wild‐type and sipA mutant strains of S. typhimurium. Time‐lapse phase‐contrast microscopy of membrane ruffle propagation in live cells confirmed that the sipA mutant induced membrane ruffles as efficiently as the wild‐type bacteria. These studies also revealed that, after ruffle propagation, individual sipA mutant S. typhimurium either invaded more slowly than wild‐type bacteria or failed to invade at all. Furthermore, although wild‐type S. typhimurium typically maintained a position central to the developing membrane ruffle, sipA mutant bacteria frequently moved initially to the periphery of the spreading ruffle and were sometimes observed to detach from it. A wild‐type pattern of invasion was restored to the sipA mutant after the introduction of sipA on a plasmid. Together, these data indicate that loss of sipA significantly decreases the efficiency of S. typhimurium invasion at the early stages of infection without affecting its ability to induce membrane ruffles. It thus appears that the secreted effector protein SipA promotes invasion by a previously unrecognized mechanism separate from the induction of membrane ruffling per se.

Real-time imaging of type III secretion: Salmonella SipA injection into host cells

“…This study presents a real-time visualization of the type III secretion process and explains why S. typhimurium can trigger host cell responses within 1 min after docking (29)(30)(31). However, the amounts and kinetics of effector protein delivery did vary somewhat between individual bacteria.…”

Molecular Pathogenesis ofSalmonella entericaSerotype Typhimurium-Induced Diarrhea

“…Inactivation of sipA drastically reduces the ability of Salmonella serotype Typhimurium to elicit fluid accumulation (Fig. 4) and an inflammatory response in bovine ligated ileal loops (115) but causes only a short (5 min) delay during invasion of epithelial cell lines in vitro (46,118). Finally, inactivation of either sopA or sopD has no effect on the ability of Salmonella serotype Typhimurium to enter epithelial cell lines but reduces its ability to elicit fluid accumulation (Fig.…”

Impact ofSalmonella entericaType III Secretion System Effectors on the Eukaryotic Host Cell

“…Salmonella invasion protein A (SipA) is a protein encoded on SPI1 [220] that is delivered into the host cell through T3SS1 [21]. This effector has a role in invasion of epithelial cells by modulating actin assembly through its Cterminal actin-binding domain, stabilizing F-actin filaments, and increasing the bundling activity of the host protein Tplastin [21,[221][222][223]. SipA contributes, together with other T3SS1 effectors, to the disruption of tight junctions, protein complexes intimately linked to the actin cytoskeleton that are located at the interface between epithelial cells [38].…”